1. Mapping Oxygen Accessibility to Ribonuclease A Using High-Resolution NMR Relaxation Spectroscopy 
Ching-Ling Teng, Robert G. Bryant 
Biophysical Journal 
Volume 86, Issue 3, Pages (March 2004)
DOI: /S (04)74240-X
Copyright © 2004 The Biophysical Society Terms and Conditions

2. Figure 1
The saturation recovery response of two representative crosspeaks, (120:HD1, HE1) and (120:HE1, HD1), in the COSY spectra for RNase A measured at pH 3.2 and 30°C. Although 120:HD1 and 120:HE1 are coupled to one another, their respective oxygen induced spin-lattice relaxation rate contributions can be measured separately in the COSY experiment. The spatial resolution of the oxygen accessibility map is limited only by the available spectroscopic resolution.
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions

3. Figure 2
The ribbon structure of ribonuclease A labeled with the structural elements. The boxed labels indicate the residues of the active site triad.
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions

4. Figure 3
(a) Color representation of the oxygen-induced proton relaxation rate constant changes (ΔR1). All spectrally resolved protons are represented as space-filling spheres that are superimposed on the ribbon structure of ribonuclease A. The color intensity changes linearly from white to pink to red in proportion to the changes of relaxation rate constants in the range from 0s−1 to 4s−1. Relaxation rate constant contributions that are larger than 4s −1 are colored as dark red. (b–f) Protons whose oxygen-induced relaxation rate constants >4s −1 are identified in red spheres but are also shown as yellow groups to facilitate discussion in the context of structural features. (b) Phe-8:HD1, Phe-8:HE1, Glu-9:HA, Leu-51:HA, Leu-51:HG, Val-54:HB, Gln-55:HB1, Pro-117:HB2, and Pro-117:HG2 are highlighted in yellow and are in a buried oxygen binding pocket. (c) Asp-83:HB1, Lys-98:HB1, Thr-100:HB, Thr-100:HG1, and Ala-102:HB1 are located at the major hairpin (β4-β5) forming a surface β groove. (d) Ser-21:HA and Ser-22:HA are located near a surface pocket between the helix linker, connecting α1 and α2, and the antiparallel β5. A cocrystallized 2-methyl-2-propanol molecule is found to bind nearby (Wlodawer et al., 1988), which suggests higher hydrophobicity. (e) Asn-27:HB1, Cys-95:HB1, and Ala-96:HB1 are located at the surface cleft between α2 and β5. Asn-113:HA is located right at the sharp turn of the β6-β7 hairpin. (f) Cys-72:HB2, Thr-70:HB, Thr-70:HG1, and Asn-71:HB2 are located at the β2-β3 hairpin region. (g–h) Protons for which the oxygen-induced relaxation rate constants are <0.6s−1 are shown as blue spheres but are also shown as yellow groups to facilitate discussion in the context of structural features. (g) Gln-11:HA, Cys-26:HA, Leu-35:HD11, Phe-46:HA, Phe-46:HB1, Phe-46:HD1, Phe-46:HE1, Phe-46:HZ, and Cys-84:HB1 are buried in the first structural lobe and are located near the central β-strand. (h) Tyr-73:HB1, Gln-74:HB1, Ile-107:HG11, Val-108:HG21, Ala-109:HA, and Cys-110:HB2 are buried in the second structural lobe and are located at the centers of the β3 and β6 strands. Ser-75:HB1, on the other hand, is located near the protein surface. However, invariant water molecules are found nearby (Sadasivan et al., 1998). The measured and computed parameters for the above high rate and low rate protons are listed in Tables 1 and 2, respectively. The structural model of RNase A (7RSA) is courtesy of the Protein Database (Wlodawer et al., 1988).
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions

5. Figure 4
Relaxed-eyed steric pairs of the computed molecular surface of ribonuclease A sensed by a surface probe of radius 1.4Å (the shorter axis of molecular oxygen) in various orientations. Those protein protons for which the oxygen-induced relaxation rate constant is >4s−1 are colored red. (a) Asp-83:HB1, Lys-98:HB1, Thr-100:HG1, and Ala-102:HB1, indicated by black arrows from left to right, are located at a rather deep surface groove between the major hairpin (β4-β5) on top of the minor hydrophobic core that connects directly to the major catalytic cleft. Blue arrow points to another surface pocket where Cys-72:HB2 is located and has large oxygen association. (b) Ser-22:HA and Ser-21:HA, indicated by black arrows from left to right, are located at a surface pocket in which a cosolute molecule, 2-methyl-2-propanol cosolvent molecule is found to bind in the x-ray structure model (Wlodawer et al., 1988).
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions

6. Figure 5
Relaxed-eyed steric pairs of the van der Waals surface of ribonuclease A. Those protein protons for which the oxygen-induced relaxation rate constant is <0.6s−1 are colored blue. (a) Gln-74:HB1 and Ile-107:HG11 (arrows) are located at a surface crevice surrounded by hydrophobic residues. An interpenetrating water network is reported in the neighborhood (Savage and Wlodawer, 1986; Wlodawer et al., 1988). (b) Ser-75:HB1 is the only surface proton with low oxygen-induced relaxation rate. Two of the invariant water molecules (W216 and W208) are found nearby (Sadasivan et al., 1998), which is consistent with the steric exclusion of oxygen caused by hydration effects.
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions

7. Figure 6
Interior protons relaxation rate differences were measured at depths >6Å and are indicated as white spheres in the ribbon representation of RNase A. Comparison with Fig. 3, g and h, shows that oxygen-induced relaxation efficiency is not simply correlated with the depth of an interior proton.
Biophysical Journal  , DOI: ( /S (04)74240-X)
Copyright © 2004 The Biophysical Society Terms and Conditions